The tumor suppressor gene p53 mediates a variety of antiproliferative responses, including cell cycle arrest, apoptosis, and senescence. The mechanism by which p53 regulates senescence is poorly understood. Sustained overexpression of wild-type p53 in tetracycline-regulated EJ-p53 bladder carcinoma cells rapidly triggered a senescence program with characteristic features and thus, represents an innovative approach to study the molecular mediators of p53-induced senescence, p53-induced senescence is characterized by a dramatic decrease in mitochondrial membrane potential (delta psi mu)) and by a small fraction of induced p53 localizing to mitochondria similar to that reported in p53-dependent apoptosis. However, unlike apoptotic cells, p53-induced = senescent cells exhibit decreased responsiveness to pharmacologic agents that activate or inhibit the mitochondrial permeability transition pore complex (PTPC), and decreased expression of mitochondrial adenine nucleotide translocase (ANT), a major component of the PTPC. We hypothesize that induction of p53 with subsequent localization of p53 to mitochondria and dissipation of delta psi mu coupled early events in cells committing to either, senescence or apoptosis, and that p53-induced senescence is mediated by structural and functional changes in the PTPC that are distinct from those associated with p53-mediated apoptosis. Based on our experimental evidence we will perform the following specific aims: 1) To analyze the structure-function relationship 9f p53 and senescence. 2) To investigate the role of p53 in direct signaling at the level of mitochondria in p53-indueed senescence. 3) To analyze the role of structural and functional changes in the mitochondrial PTPC in mediating p53-induced senescence. It is critical to understand the molecular basis for p53-induced senescence as it represents an alternative mechanism for a tumor cell to permanently exit the cell cycle and therefore may be the basis for alternative cancer therapy.